Carotenoids are natural pigments that are useful as feed additives, food additives, pharmaceutical agents, and the like. Examples of carotenoids include astaxanthin, canthaxanthin, zeaxanthin, β-cryptoxanthin, lycopene, β-carotene, phoenicoxanthin, adonixanthin, echinenone, asteroidenone, and 3-hydroxyechinenone.
Among carotenoids, astaxanthin is useful as feed additives, for example, as a body color-improving agent for cultivated fishes such as salmon, trout, and sea bream or an egg yolk color-improving agent for poultry. Moreover, astaxanthin is highly valuable in the industries as safe natural food additives and health food materials.
Similar to astaxanthin, adonixanthin and phoenicoxanthin are also expected to be used as feed additives, food additives, pharmaceutical agents, and the like once industrial production methods thereof are established. Furthermore, β-carotene is used as feed additives, food additives, pharmaceutical agents, and the like; canthaxanthin is used as feed additives, food additives, cosmetics, and the like; and zeaxanthin is used as food additives, feed additives, and the like. In addition, other carotenoids such as lycopene, echinenone, β-cryptoxanthin, 3-hydroxyechinenone, and asteroidenone are also expected to be used as feed additives, food materials, and the like. Known methods for producing these carotenoids include chemical synthesis methods, methods of extraction from natural sources, and production methods using microorganisms.
As methods for chemically synthesizing astaxanthin, a method utilizing conversion of β-carotene (Non-Patent Document 1) and a method employing synthesis from C15 phosphonium salt (Non-Patent Document 2) are known. Astaxanthin produced by such chemical synthesis methods is commercially available as a feed additive. In addition, since astaxanthin is present in fishes such as sea bream and salmon as well as crustaceans such as shrimp, crab, and krill, it may also be extracted therefrom.
Methods that have been reported for producing astaxanthin using microorganisms include a culture method using green alga Haematococcus pluvialis (Patent Document 1), a fermentation method using red yeast Phaffia rhodozyma (Patent Document 2), and a fermentation method using a bacterium belonging to the genus Paracoccus (hereinafter occasionally referred to as a “Paracoccus bacterium”).
Examples of astaxanthin-producing Paracoccus bacteria include strains E-396 and A-581-1 (Patent Document 3 and Non-Patent Document 3). Examples of other astaxanthin-producing Paracoccus bacteria include Paracoccus marcusii strain MH1 (Patent Document 4), Paracoccus haeundaensis strain BC74171 (Non-Patent Document 4), Paracoccus bacterial strain N-81106 (Patent Document 5), and Paracoccus sp. strain PC-1 (Patent Document 6).
There have been several problems concerning the above-mentioned carotenoid production methods. For example, chemical synthesis methods have unfavorable impression on consumers from a safety perspective. Extraction from natural sources such as shrimp and crab is associated with high production costs. In addition, production using a green alga or yeast results in low productivity and has difficulty in extraction of a carotenoid therefrom due to strong cell walls thereof.
Meanwhile, bacteria belonging to the genus Paracoccus are advantageous in that proliferation rates thereof are fast, carotenoid productivities thereof are high, and carotenoids can readily be extracted therefrom, etc. Several methods for culturing such bacteria have been reported. For example, Patent Document 7 discloses a method characterized by adding an iron salt during culture. Patent Document 8 discloses a method characterized by restricting the carbon source concentration. However, such culture methods are problematic in that large amounts of canthaxanthin accumulate during production of astaxanthin.
Canthaxanthin is a useful feed additive for improving the color tone of salmon meat or hen egg yolk, while the ADI (acceptable daily intake) thereof is limited to 0.03 mg/kg of body weight and the upper limits of the amounts of canthaxanthin acceptable to be added to feed are stipulated as 25 mg/kg and 8 mg/kg for salmon and laying hens, respectively, in Europe (Non-Patent Document 5). Thus, it is necessary to control the canthaxanthin content at a low level when producing astaxanthin using a microorganism. Patent Document 9 discloses a method wherein canthaxanthin production is reduced by controlling the dissolved oxygen concentration. However, such a method also results in significant reduction in the concentration of produced astaxanthin and thus is not practical in terms of production cost.